PROJECT SUMMARY The goal of this project is to develop an epigenetic microarray platform (EpiSeraTM) as a diagnostic and staging tool for autoimmune diseases. Autoantibodies directed against nuclear antigens, including histone proteins and double-stranded DNA (dsDNA), are prevalent in serum of patients with systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and other autoimmune diseases. Assays to detect circulating autoantibodies toward histones and/or dsDNA are commonly used to help diagnose and stage SLE, and assays capturing autoantibodies with citrullinated histone peptide antigens are highly predictive and selective for the diagnosis of RA. However, these tests fail to detect disease in all patients and poorly reflect the heterogeneous nature of these complex pathologies. The development of new autoimmune biomarkers are desperately needed and will enable improved personalized treatment strategies as well as provide clinicians with indicators to better track disease progression and therapy response. Nucleosomes are heavily decorated with histone post-translational modifications (PTMs) and DNA methylation. We hypothesize that capturing plasma autoantibodies with histone and DNA antigens harboring defined modifications has clinical potential for autoimmune disease diagnosis, staging, and treatment response. Preliminary data suggests that serum from SLE patients contains autoantibodies specific for unique sets of histone PTMs. The innovation of this project is the development of a microarray platform to profile a patients? autoantibody epigenetic footprint (comprised of both DNA and histone modifications) as a next-generation diagnostic assay. For proof of concept, we will focus on patients with SLE and RA; these pathologies are strongly associated with high levels of nuclear targeting autoantibodies. Here, we will develop EpiSera, a new specialized microarray platform that includes >150 unique modified peptides and DNA oligos covering major nucleosome modifications for the rapid capture and characterization of human immunoglobulin (Ig) isotypes (Aim 1). EpiSera will contain 12 subarrays, enabling users to probe multiple patient samples / dilutions per array. We will use EpiSera to profile autoantibodies from patients with SLE and RA (Aim 2). This project will be considered successfully complete when EpiSera can be used to distinguish SLE and RA patients from each other and from healthy controls. Our study has the potential to develop a robust clinical tool to identify new biomarkers for SLE / RA diagnosis, staging, and therapeutic response. In Phase II, we will expand the scope of our preclinical studies to better define key epigenetic profiles that delineate SLE and RA patients based on disease severity, progression, etc. At the conclusion of Phase II, we will begin to market EpiSera for pre-clinical and research applications and be well-positioned to initiate and receive FDA 510(k) clearance.